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Uterine fibroids (UF), also called leiomyomas, are observed in nearly 70% of women of reproductive age. These tumors disrupt normal uterine function and cause significant physical and psychological health problems. Although heritable genetics is a significant risk factor, specific genetic variations and gene targets causally associated with UL are poorly understood. We performed a meta-analysis on existing fibroid genome-wide association studies (GWAS) and integrated the identified risk loci and potentially causal single nucleotide polymorphisms (SNPs) with epigenomics (H3K27me3), transcriptomics (RNA-Seq), 3D chromatin organization (Hi-C) and single-cell atlas (scRNA-Seq) of primary samples from UF patients. The integrative analysis identified 24 risk loci and 394 potential gene targets associated with UF. Among these, 106 of them were differentially expressed in UF tumors. Critically, the integrative analysis with single-cell RNA sequencing revealed the cell types with aberrant expression of these target genes in normal and UF tissue. Finally, CRISPR-based epigenetic manipulation (dCas9-KRAB) of two risk loci in a disease-relevant cell type fine-mapped the gene targets. These findings and the methodological approach indicate the effectiveness of integrative multi-omics data combined with locus-specific epigenetic editing approaches to identify disease-relevant risk loci and their target genes.

Pancreatic ductal adenocarcinoma (PDA) cells reprogram their transcriptional and metabolic programs to survive the nutrient-poor tumor microenvironment. Through in vivo CRISPR screening, we discovered islet-2 (ISL2) as a candidate tumor suppressor that modulates aggressive PDA growth. Notably, ISL2, a nuclear and chromatin-associated transcription factor, is epigenetically silenced in PDA tumors and high promoter DNA methylation or its reduced expression correlates with poor patient survival. The exogenous ISL2 expression or CRISPR-mediated upregulation of the endogenous loci reduces cell proliferation. Mechanistically, ISL2 regulates the expression of metabolic genes, and its depletion increases oxidative phosphorylation (OXPHOS). As such, ISL2-depleted human PDA cells are sensitive to the inhibitors of mitochondrial complex I in vitro and in vivo. Spatial transcriptomic analysis shows heterogeneous intratumoral ISL2 expression, which correlates with the expression of critical metabolic genes. These findings nominate ISL2 as a putative tumor suppressor whose inactivation leads to increased mitochondrial metabolism that may be exploitable therapeutically.

Aging occurs due to a combination of several factors, such as telomere attrition, cellular senescence, and stem cell exhaustion. The telomere attrition-dependent cellular senescence is regulated by increased levels of SMAD specific E3 ubiquitin protein ligase 2 (smurf2). With age smurf2 expression increases and Smurf2 protein interacts with several regulatory proteins including, Smad7, Ep300, Yy1, Sirt1, Mdm2, and Tp53, likely affecting its function related to cellular aging. The current study aimed at analyzing smurf2 expression in the aged brain because of its potential regulatory roles in the cellular aging process. Zebrafish were used because like humans they age gradually and their genome has 70% similarity. In the current study, we demonstrated that smurf2 gene and protein expression levels altered in a region-specific manner during the aging process. Also, in both young and old brains, Smurf2 protein was enriched in the cytosol. These results imply that during aging Smurf2 is regulated by several mechanisms including post-translational modifications (PTMs) and complex formation. Also, the expression levels of its interacting partners defined by the STRING database, tp53, mdm2, ep300a, yy1a, smad7, and sirt1, were analyzed. Multivariate analysis indicated that smurf2, ep300a, and sirt1, whose proteins regulate ubiquitination, acetylation, and deacetylation of target proteins including Smad7 and Tp53, showed age- and brain region-dependent patterns. Our data suggest a likely balance between Smurf2- and Mdm2-mediated ubiquitination, and Ep300a-mediated acetylation/Sirt1-mediated deacetylation, which most possibly affects the functionality of other interacting partners in regulating cellular and synaptic aging and ultimately cognitive dysfunction. (C) 2020 IBRO. Published by Elsevier Ltd. All rights reserved.